PROJECT SUMMARY/ABSTRACT The expression of GnRHRs is a critical, rate-limiting step in the reproductive process, however little is known about the mechanism behind the regulation of translation of GnRHR proteins. Gonadotrope functions may be limited in times of food deprivation, which is signaled by leptin deficiency, however there is a fundamental gap in understanding how leptin regulates gonadotropes. Leptin's importance to gonadotropes is highlighted by the infertile mouse model in which all isoforms of leptin receptors (LEPR) on gonadotropes are ablated, and the fact that the mutant gonadotropes have severely reduced GnRHR protein, but not mRNA levels. These findings provide important clues as to how leptin links fertility to metabolic status, however the molecular mechanisms underlying leptin control of gonadotropes are unknown. Our long-term goal is to fill this knowledge gap by identifying underlying mechanisms behind the infertility in the gonadotrope Lepr-null mutant mice. The central hypothesis to be tested is that loss of leptin signaling in gonadotropes prevents the normal diestrous upregulation of GnRH receptors causing a blunted or absent LH surge and infertility. A secondary hypothesis that leptin acts through posttranscriptional mechanisms to optimize gonadotrope function. The proposed studies will focus on three specific aims. Specific Aim 1 will determine if loss of leptin signaling in gonadotropes reduces fertility through prevention of the diestrous upregulation of GnRHR protein levels. These studies will ascertain if the Lepr-null gonadotropes are held in persistent diestrus because of low GnRHR proteins and if the normal surge in serum gonadotropins is absent. The efficacy of exogenous GnRH in the rescue of GnRHR levels in mutants and restoration of cyclicity and fertility will be tested. Specific Aim 2 studies will determine if leptin signaling to mRNA translational control mechanisms is required for gonadotrope function. These studies will globally assess leptin regulation of gonadotrope mRNA translation and specifically address control of Gnrhr mRNA translation. Genetic knockdown of identified candidate translational repressors will be used to restore GnRHR expression in mutant gonadotropes. Specific Aim 3 studies will determine if food deprivation recapitulates the effect of loss of leptin signaling to gonadotropes, including repression of Gnrhr mRNA translation. These in vivo studies will test the hypothesis that loss of leptin signaling during food deprivation also prevents diestrous upregulation of GnRHR. The studies will also determine if the in vitro deletion of identified translational control mechanisms (e.g. MSI, miRNAs) alleviates the fasting-induced repression of Gnrhr mRNA translation. Our research is innovative because it investigates the novel concept that leptin may regulate gonadotrope function at post-transcriptional levels. These findings are significant because clinical protocols used in infertility treatments (e.g. clomiphene) depend on normal responses to endogenous GnRH, which we have shown depends on normal leptin signaling.